Enrichment and tagging of glycosylated proteins

Abstract

A method useful in the analysis of glycosylated proteins, in which a mixture containing glycosylated proteins and unglycosylated proteins is contacted with a resin that includes a nucleophile bound to a solid support via a linker. The contacting is performed under conditions sufficient to result in removal of the glycosyl group from the glycosylated proteins and to concomitantly result in the deglycosylated proteins covalently bound to the solid support. The deglycosylated proteins bound to the solid support may be rinsed to remove proteins that are not covalently bound to the solid support. The deglycosylated proteins are released from the solid support and may be subjected to further purification and / or analysis.

Description

FIELD OF THE INVENTION[0001]The invention relates generally to analysis of proteins and peptides. More specifically, the invention relates to solid phase methods for enrichment of glycosylated proteins and / or peptides. Such methods are useful for preparing the glycosylated proteins and / or peptides for further analysis.BACKGROUND OF THE INVENTION[0002]Sequencing of the human genome, and the genomes of other species, has emphasized the fact that the expression and properties of a protein are often dependent on posttranslational modifications and, thus, cannot be predicted from the DNA sequence. This realization has spurred an interest in proteomics, the study of protein expression within a cell under defined conditions.[0003]Traditionally, proteins from biological samples have been isolated and identified by separating the proteins using 2-D gel electrophoresis followed by identification of the protein using mass spectrometry. However, this method is time consuming and can only detect...

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Problems solved by technology

However, this method is time consuming and can only detect proteins that are highly abundant in the biological sample.

Severe streaking causes deterioration in resolution of the electrophoretic separation when high loading is used in an attempt to visualize less abundant proteins.

Particular difficulties have been encountered in attempts to use 2-D gel electrophoresis / mass spectrometry to study glycosylated proteins, as they are often present in low abundance.

While the above methods have greatly facilitated the study of glycoproteins, they have various disadvantages, such as requiring multiple purification or clean-up steps in isolating or labeling a target protein or collection of proteins.

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